Serum Exosomal MRPS33 MRNA As A Novel Marker For Hepatocellular Carcinoma

Objective To explore whether mitochondrial ribosomal protein S33 messager RNA(MRPS33 mRNA)could be used as a diagnostic and prognostic marker for hepatocellular carcinoma(HCC).Methods The serum differential exosomal mRNAs were screened by RNA deep sequencing(RNA-seq),in which MRPS33 mRNA was the remarkerbly high expressed and candicated as the diagnostic indicator for HCC.First,quantitative reverse transcription polymerase chain reaction(qRT-PCR)was applicated to detect the relative expression levels of exosomal MRPS33 mRNA in the culture supernatant from 2 human hepatoma cell lines and 1 normal hepatocyte cell line.Meanwhile,a test set including 30 HCC patients,30 chronic hepatitis B(CHB)patients and 30 healthy subjects was enrolled,the serum levels of exosomal MRPS33 mRNA from those were measured by qRT-PCR.Second,80 HCC patients,82 liver cirrhosis patients(LC),65 CHB patients and 60 healthy subjects were recruited as an independent set to validate the power of serum exosomal MRPS33 mRNA discriminating HCC from CHB.In further,the receiver operating characteristic(ROC)curves were constructed and the area under ROC curves(AUCs)were obtained to assess the diagnostic efficacy of MRPS33 mRNA alone or/and in combination with alpha-fetoprotein(AFP)in HCC.Finally,the relationship between MRPS33 mRNA expression and clinicopathological characteristics and overall survival(OS)was analyzed in 80 HCC patients.Results A total of 220 serum exosomal mRNAs with differential expression in HCC were screened by RNA-seq,and MRPS33 mRNA was significantly up-regulated(6.85-fold change)in HCC patients.The expression levels of MRPS33 mRNA in 2 culture medium exosomes isolated from 2 hepatoma cell lines(SK-HEP-1 and BEL-7402)and 1 normal liver cell line(LO2)were significantly different(all P<0.05).The difference in the expression levels of serum exosomal MRPS33 mRNA among HCC patients,CHB patients and healthy controls were significantly observed(all P<0.05).The level of serum exosomal MRPS33 mRNA in patients with HCC was significantly higher than that in LC group,CHB group and healthy subjects(all P<0.05).Similary,the level of MRPS33 mRNA in LC was significantly higher than that in CHB patients and healthy subjects(All P<0.05).When the MRPS33 mRNA cut-off value was 0.249 for identifying HCC from CHB,yielding AUC value of 0.823,with a sensitivity of 83.8%and a specificity of 77.0%.Furthermore,a assay in combination exosomal MRPS33 mRNA and AFP obtained AUC of 0.907,with a sensitivity 82.5%and a specificity 88.0%.In addition,there were no significant correlation between MRPS33 mRNA expression and gender,age,serum HBsAg status,tumor size,TNM stage and cirrhosis with HCC(all P>0.05),whereas the relationship were found in respect to Child-Pugh classification,portal vein tumor thrombus,lymph node metastasis and OS(all P<0.05).Conclusion Serum exosomal MRPS33 mRNA is a promising diagnostic and prognostic marker for HCC.In the meantime,an assay in combination of serum exosomal MRPS33 mRNA and serum AFP could improve the diagnostic efficacy for HCC.